2-hydroxy-pteroic acid and the n-formylated derivatives thereof



Paten ted Dec. 25, 1951 UNITED STATES PATENT OFFICE Z HYDROXY-S'ZZL ZICACID AND I i I I N THE N- FORMYLATED DERIVA- TIVES THEREOF Donald E.Wolf, Franklin Township, Union County, N. J., assignor to Merck & 00.,Inc., Rahway, N. J a corporation of New Jersey N Drawing.

Original application August 10,

1946, Serial No. 689,815. Divided and this application March '22, 1947,Serial No. 736,585

Claims.

- on R N q -M3 110- J \N N/ wherein R represents hydrogenand formylsubstituents.

A compound of this general structure wherein R isa formyl group is fullydefined by the chemical name 4- (2,4-dihydroxy-6-pteridyl)methyl]formylamino)benzoic acid and will hereinafter he referred to asdesiminorhizopterin.

A compound of the above formula wherein R is hydrogen is fully definedby the chemical name 4-( (2,4 dihydroxy 6 pteridyl) methyl]amino)benz0ic acid and will hereinafter be referred to asdesiminoaporhizopterin.

In accordance with my invention these new compounds may be prepared byreacting nitrous acid with rhizopterin 4-([(2-amino4-hydroxy- 6pteridyl)methyllformylamino)benzoic acid) to form desiminorhizopterin(4- E (2,4-dihydroxy- 6 pteridyl-)methyllformylamino) benzoic acid) andhydrolyzing the latter compound to form desiminoaporhizopterin (4 ([(2,4dihydroxy-B- pteridyl) methyllaminmbenzoic acid). This reaction can beindicated graphically as follows:

GOOH

The starting material, 4-([(2 amino 4 hydroxy 6 pteridyl)methyllformylamino) benzoic acid, commonly referred to'as rhizopterin,is the pterin-like acidic chemical compound obtained from fumaric acidfermentation liquor. This general process and rhizopterin therebysecured is disclosed and claimed in a copending application of mycolleagues, Keresztesy and Rickes, Serial No. 536,434, filed May 19,1944, now Patent No. 2,478,404.

Desiminorhizopterin is obtained in accordance with this invention byreacting an acid solution of rhizopterin with a mixture capable ofevolving nitrous acid. Thus, when mixtures such as sodium nitrite andacetic acid are added to a concentrated hydrochloric acid solution ofrhizopterin, nitrous acid is evolved in-the reaction mixture and reactsupon the rhizopterin to form desiminorhizopterin. The reaction ispreferably conducted at room temperature at which temperature thereaction is completed in about onehour. The solution is concentrated todryness in vacuo and the residue extracted with water to removeinorganic salts. The crude reaction compound is P thereby'recovered as ayellow solid which can be purified by recrystallization from hot water.

Desiminoaporhizopterin can be prepared by reacting desiminorhizopterinwith a concentrated acid, diluting the reaction mixture with water andrecovering the yellow precipitate thus formed. The crudedesiminoaporhizopterin is purified by dissolving it in ammoniumhydroxide and acidifying the solution slowly with acetic acid thuscausing separation of the product as a crystalline precipitate.

The following example illustrates a method of carrying out the presentinvention, but it is. to

of illustration and not of limitation.

Example 26 mg. of rhizopterin are dissolved in cold contratedhydrochloric acid and a mixture of sodium nitrite and acetic acid isadded. The reaction is allowed to proceed for 1 hour at roomtemperature, after which the solution is evaporated to dryness in vacuo.The residue is extracted with cold water to remove inorganic salts, andthe the insoluble desiminorhizopterin, i. e. 4-(2,4-dihydroxy-6-pteridyl) methyl] formylamino) benzoic acid, remains asa light yellow solid. It is purified by recrystallization from hotwater. The pure product melts at 319-322".

Anal.-Ca1cd. for CisHuNsOs: C, 52.79; H, 3.25, Found: C, 52.62; H, 2.93.C, 52.78; H, 3.36.

2 mg. of desiminorhizopterin are suspended in strong hydrochloric acid(about 7 to 8 N solution), and the suspension heated in the steam bathforminutes Dilution with water causes the precipitation of a brightyellow desiminoaporhizopterin i. e., 4-([(2,4-dihydroxy-6-pteridyl)-methyllamino)benzoic acid, which is centrifuged and washed with water.The. crude desiminoaporhizopterin is purified by dissolving it in diluteammonium hydroxide and slowly acidifying the solution with acetic acid.Pure desiminoaporhizopterin is thus obtained in the form of fine yellowcrystals.

AnaL-Calcd. for C14H11N5O42 c, 53.67; H, 3.54. Found: C, 53.83; H,3.52'.

Pure desiminoaporhizopterin 'is soluble in alkalies only, and ischaracterized by the following ultra-violet absorption spectrum:

' v Wave lengths in A. E%'

pH: 11. 2780 92c wherein R'is selected from the class consisting ofhydrogen and formyl substituents. I 2. 4 ([(2,4 dihydroxy 6 pteridyl)methyl] f0rmy1amino)benzo-ic acid.

3. '4-([ (2,4 dihydroxy 6 pteridyl) methyl]- amino)benzoic acid. 4. The;process that comprises reacting a mineral acid solution of4-([(2-amino-4-hydroxy-6- pteridyl) methyl] formylamino) benzoic acidwith nitrous acid, recovering 4-([(2,4-dihydroxy-6- pteridyl) methyl]formylamino) benzoic acid thus formed, hydrolyzing the latter compoundand recovering 4'- (2,4-dihydroxy-6-pteridyl)' methyl] amino)benzoicacid.

5. The process that comprises reacting a mineral acidsolution of*l-(E(2-amino-4-hydroxy-6- pteridyl) methyl] formylamino) benzoic acidwith a mixture of sodium nitrite and acetic acid, evaporating themixture to dryness, reacting the residue formed with water toprecipitate 4- 2,4 dihydroxy 6 pteridyhmethyllfor- 4 my1amino)benzoicacid, recovering said precipitate, reacting the precipitate withhydrochloric acid, diluting said acid mixture with water to precipitate4 ([(2,4 dihydroxy 6 pteridyl) methy1]amino)benzoic acid and recoveringthe latter compound. 7

6. The process that comprises reacting a mineral acid solution of4-([(2*amino-4-hydroxy-6- pteridyl)methyllformylamino)benzoic acid withnitrous acid .and recovering l-([(2,4-dihydroxy- G-pteridyl) methyl]formylamino) benzoic acid.

7. The process that comprises dissolving 4- (E (2 amino 1 4 hydroxy 6pteridyl) methyl] formylarnincbbenzoic acid in hydrochloric acid,reacting the acid solution thus formed with a mixture of sodium nitriteand acetic acid and recovering 4- (2,4-dihydroXy-6-pteridyl) methyl]formylamino benzoic acid.

8. The process that comprises reacting a mineral acid solution of4-([(2-amino-4-hydroxy-6- pteridyl) methyllformylamino)benzoic acid witha mixture of sodium nitrite and acetic acid evaporating the mixture todryness, reacting the residue formed with water to precipitate 4- ([(2,4dihydroxy 6 pteridyDmethyllformylamino) benzoic acid and recovering thelatter compound. 7

9. The process that comprises hydrolyzing 4- ([(2,4 dihydroxy 6pteridyl)rnethyl]for-'- mylamino)benzoic acid and recovering 4 (["(2,4dihydroxy 6 pteridyl)methyl]amino)benzolc acid.

10. The process that comprises reacting 4- ([(2,4 dihydroxy 6pteridyl)methy1]formylamino)benzoic acid with hydrochloric acid,diluting said acid mixture with water to precipitate 4 ([(ZA dihydroxy6- pteridyl)methyl]-' amino)benzoic acid and recovering the lattercompounds. I

DONALD'E. WOLF.

REFERENCES CITED The following references are of record in the file ofthis patent:

UNITED STATES PATENTS Date Angier June 8-, 1948 OTHER REFERENCESScience, May 31, 1946, pages 667-669.

Chemical Abstracts, vol. 39, 4684 Fieser and Fieser: Organic Chemistry,"page.

Fieser and 'Fieser: Organic Chemistry, pp,-

' 235-236, 1944 edition, 1). C. Heath and Co.

1. A COMPOUND REPRESENTED BY THE FORMULA
 4. THE PROCESS THAT COMPRISESREACTING A MINERAL ACID SOLUTION OF4-(1(2-AMINO-4-HYDROXY-6PTERIDYL)METHYL)FORMYLAMINO) BENZOIC ACID WITHNITROUS ACID, RECOVERING4-($(2,4-DIHYDROXY-6PTERIDYL)METHYL)FORMYLAMINO) BENZOIC ACID THUSFORMED, HYDROLYZING THE LATTER COMPOUND AND RECOVERING4-($(2,4-DIHYDROXY-6-PTERIDYL)METHYL)AMINO)BENZOIC ACID.